We have identified a new class of DNA sequences that are found only in heterochromatin, yet' differ significantly from the tandemly repeated DNAs previously characterized as heterochromatin-specific sequences. This new class of sequences, the HeT DNA family, is from Drosophila melanogaster. The polytene chromosomes plus the many chromosomal rearrangements and marker mutations that are available make D. melanogaster the best model system for studying the structure and function of heterochromatin; however we believe that the results of these studies will give insights into the chromosomes of all metazoa. A major component of HeT DNA is a transposable element named HeT-A. Although HeT-A elements are associated with sequences that are shared between telomeric and pericentric heterochromatin (sequences that make up other components of HeT DNA), HeT-A elements themselves are found only in telomeric heterochromatin. HeT-A elements have been added to the broken ends of the ten "healed" terminal deletions now known in D. melanogaster. The elements have apparently been added by transposition events involving no sequence homology, instead the structure of the junction with the broken chromosome suggests retrotransposition. If this is true, the retrotransposition is remarkably restricted.- HeT-A elements differ from the known transposable elements in having >2 kb of conserved short-range sequence repeats on one end (the end that invariably is attached to the broken chromosome end). HeT-A elements are abundant on the telomeres of unbroken chromosomes. These features of HeT-A elements suggest that, although they are transposable elements, they may have a structural role in telomere organization or maintenance. HeT-A elements do not resemble the very short repeats found at the telomeres in other organisms but they appear to be similar to the "telomere-associated" sequences found in metazoan chromosomes. Thus other organisms may also have transposable elements that are not "selfish DNA". We propose to characterize the protein encoded by HeT-A and any RNAs transcribed from the element. There is reason to think that transposition is inducible. We will seek to determine in what cell types and under what conditions RNA and protein are expressed as a step toward understanding the induction of transposition. We will seek HeT-A elements in a second Drosophila species to analyze the evolutionarily conserved features of the element. We propose several transgenic experiments to test conclusions about possible functions of the protein encoded by HeT-A and of the non-coding repeated segment. Finally, we will attempt to use the sequences that we define to construct functional Drosophila mini- chromosomes. These chromosomes will also allow direct tests of conclusions.